DESCRIPTION Calcium-regulated secretion of neurotransmitters is the primary mode of communication in the nervous system. Neurotransmitter secretion is mediated by a unique organelle, the synaptic vesicle, which undergoes repetitive rounds of filling, targeting, priming, exocytosis and endocytosis in the presynaptic terminal. Several of these stages are accomplished via the sequential formation and dissociation of protein complexes. Identifying the protein complexes involved, determining where in the vesicle cycle each complex acts and describing how calcium regulates their action is therefore central to understanding the molecular mechanisms of neurotransmitter release. The applicant and her students have identified a protein complex that contains the synaptic vesicle proteins SV2 and synaptotagmin. SV2 is a transporter-like membrane glycoprotein hypothesized to function either as a small molecule transporter or as a component of the fusion pore that initiates transmitter release. Synaptotagmin is a calcium-binding protein required for normal calcium-regulated exocytosis. The presence of these proteins in all synapses, together with the observation that calcium regulates their interaction, suggests that the SV2 synaptotagmin complex plays a crucial role in neurotransmitter secretion. To identify that role the applicant proposes four aims designed to determine the physiological function of the complex and how that function is regulated. 1. The applicant will determine the components of the SV2 synaptotagmin complex. 2. The applicant will test the effect of disruption of SV2-synaptotagmin interaction on regulated secretion in PC12 cells. 3. The applicant will test whether all eight isoforms of synaptotagmin demonstrate calcium-dependent interactions with SV2. 4. The applicant will determine whether phosphorylation regulates the interaction of SV2 and synaptotagmin. These studies will provide basic information required to understand the changes in synaptic efficacy that underlie both learning and memory as well as multiple neuropathologies.